Glycosphingolipids (GSL)-enriched 'rafts'and caveolae are membrane microdomains that putatively function as lateral organizing sites for signaling proteins involved in oncogenesis. Because the overexpression of select caveolar proteins, i.e. caveolin-1, is associated with tumor cell survival, aggression and metastatic potential, targeting GSL-enriched caveolae may prove useful as a new therapy for the functional disruption of metastatis, tumorigenesis, and tumor progression. The processes by which GSL-enriched domains are formed and maintained are not well defined but are expected to involve specific proteins that can bind and transfer GSLs between and within cells. Our objective is to elucidate the structure of human glycolipid transfer protein (GLTP) and related orthologs and to identify/-characterize folding domains responsible for glycolipid liganding selectivity using structural, dynamical, and mutational approaches by taking advantage of the structural expertise of the Dinshaw Patel (Sloan-Kettering Institute, NY) and the molecular biological and glycosphingolipid expertise of the Rhoderick Brown (Hormel Institute, MN) laboratories. The rationale for the research is that, solving the structure of GLTP and related orthologs in their apo and glycolipid-liganded states will enable mapping of the protein domains and associated key amino acid residues involved in GLTP functionality. Acquiring this knowledge will provide a foundation for pursuing the future development of pharmacologic agents that can specifically target GLTP in oncogenic cells displaying aberrant GLTP activity. The proposed work is innovative because it capitalizes on the first-ever, structural insights into human GLTP. The novel two-layer, all alpha helical topology of GLTP differs distinctly from the folding topologies of other known lipid binding/transfer proteins, suggesting that the GLTP folding motif defines a novel family of proteins. The studies will take advantage of our recent successes in the molecular cloning and expression of GLTP and related point mutants. It is our expectation that the proposed structural-dynamics-mutational studies of human GLTPs and related orthologs will provide unparalleled insights into the functional workings of this emerging new protein family. This new knowledge is expected to be significant by providing a foundation for using GLTP in new and innovative ways, such as introducing specific GSL antigens into cancer cells to help achieve targeted destruction of diseased cells via immunotherapeutic means.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA121493-05
Application #
7807938
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Knowlton, John R
Project Start
2006-06-01
Project End
2012-04-30
Budget Start
2010-05-01
Budget End
2012-04-30
Support Year
5
Fiscal Year
2010
Total Cost
$229,987
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
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Simanshu, Dhirendra K; Zhai, Xiuhong; Munch, David et al. (2014) Arabidopsis accelerated cell death 11, ACD11, is a ceramide-1-phosphate transfer protein and intermediary regulator of phytoceramide levels. Cell Rep 6:388-99
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Samygina, Valeria R; Ochoa-Lizarralde, Borja; Popov, Alexander N et al. (2013) Structural insights into lipid-dependent reversible dimerization of human GLTP. Acta Crystallogr D Biol Crystallogr 69:603-16
Kamlekar, Ravi Kanth; Simanshu, Dhirendra K; Gao, Yong-guang et al. (2013) The glycolipid transfer protein (GLTP) domain of phosphoinositol 4-phosphate adaptor protein-2 (FAPP2): structure drives preference for simple neutral glycosphingolipids. Biochim Biophys Acta 1831:417-27
Gao, Yongguang; Chung, Taeowan; Zou, Xianqiong et al. (2011) Human glycolipid transfer protein (GLTP) expression modulates cell shape. PLoS One 6:e19990
Kenoth, Roopa; Kamlekar, Ravi Kanth; Simanshu, Dhirendra K et al. (2011) Conformational folding and stability of the HET-C2 glycolipid transfer protein fold: does a molten globule-like state regulate activity? Biochemistry 50:5163-71
Samygina, Valeria R; Popov, Alexander N; Cabo-Bilbao, Aintzane et al. (2011) Enhanced selectivity for sulfatide by engineered human glycolipid transfer protein. Structure 19:1644-54
Kamlekar, Ravi Kanth; Gao, Yongguang; Kenoth, Roopa et al. (2010) Human GLTP: Three distinct functions for the three tryptophans in a novel peripheral amphitropic fold. Biophys J 99:2626-35
Kenoth, Roopa; Simanshu, Dhirendra K; Kamlekar, Ravi Kanth et al. (2010) Structural determination and tryptophan fluorescence of heterokaryon incompatibility C2 protein (HET-C2), a fungal glycolipid transfer protein (GLTP), provide novel insights into glycolipid specificity and membrane interaction by the GLTP fold. J Biol Chem 285:13066-78

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